The Effect of Triplet States on Non-Photochemical Hole Burning in Cytochrome b6f and Modified LH2 Complex

The Effect of Triplet States on Non-Photochemical Hole Burning in Cytochrome b6f and Modified LH2 Complex Alexandra Trempe Photosynthesis is an extraordinary process responsible for all life on Earth. In better understanding how photosynthetic complexes interact with light, we open up opportunities for advancement in environment research. While at times functionally and structurally similar, it has been shown that oxygenic photosynthetic complexes and anoxygenic photosynthetic complexes in purple bacteria possess differences. For example, oxygenic photosynthetic complexes utilise chlorophyll pigment molecules, which absorb in the red and blue regions of visible light. Purple bacteria host bacteriochlorophyll pigment molecules, absorbing at wavelengths greater than 800 nm. Important information about the functioning of photosynthetic complexes can be obtained from zero-phonon lines and phonon sidebands of the pigments. These can be observed through spectral techniques such as non-photochemical hole burning (NPHB), a type of spectral hole burning. NPHB can be explained using a two-level system, or a double-well potential, which is the simplest manner to describe the protein/pigment energy landscape. When describing NPHB, the model normally considers both a singlet ground state and a singlet excited state of the pigment. In this thesis, we will explore the limitations of only considering these two electronic state for cytochrome b6f and a modified purple bacteria light-harvesting complex II (LH2) containing chlorophylls. In particular, we originally set out to observe the illumination-intensity dependence (previously seen in cytochrome b6f) in our modified LH2 in an attempt to validate the presence of local heating of the protein by the laser beam causing the NPHB. Unfortunately, we were unable to see local heating. This brought the introduction of a triplet state to explain hole burn depth discrepancies between hole growth kinetic curves and post-burn hole spectra. Our modelling allowed us to see that transient holes caused by triplet states contributed to slower burning of persistent holes with increased intensity, as we had observed. We also examined the case of increased triplet lifetime expected when cytochrome b6f and modified LH2 were placed in a deuterated solvent. An expected triplet lifetime for both complexes in a deuterated solution is around 2 ms, while we examined triplet lifetimes ranging from 1 ms up to 5 ms. Even with a five-fold increase of the simulated triplet lifetime, it did not quite match the experimental difference between protonated and deuterated samples. Likely local heating along with triplet states explain the difference. Finally, we introduce preliminary data relating to pigment participation in the two-level system. Thermocycling hole recovery results revealed a similar recovery rate for chlorophylls placed in either B800 bacteriochlorophyll pockets or somewhere else in modified LH2. However, in both cases, the holes recovered more quickly than in the regular, bacteriochlorophyll-containing LH2. Given that the two-level system is usually expected to represent the protein environment, chlorophylls in the same location as bacteriochlorophylls were expected to recover similarly. Instead, chlorophylls in different environments recovered similarly, leading to the potential assumption that pigments also contribute to the generalised coordinate of the two-level system